Abstract
Nonribosomal peptides represent a large class of metabolites with pharmaceutical relevance. Pteridines, such as pterins, folates, and flavins, are heterocyclic metabolites that often serve as redox-active cofactors. The biosynthetic machineries for construction of these distinct classes of small molecules operate independently in the cell. Here, we discovered an unprecedented nonribosomal peptide synthetase-like-pteridine synthase hybrid biosynthetic gene cluster in Photorhabdus luminescens using genome synteny analysis. P. luminescens is a Gammaproteobacterium that undergoes phenotypic variation and can have both pathogenic and mutualistic roles. Through extensive gene deletion, pathway-targeted molecular networking, quantitative proteomic analysis, and NMR, we show that the genetic locus affects the regulation of quorum sensing and secondary metabolic enzymes and encodes new pteridine metabolites functionalized with cis-amide acyl-side chains, termed pepteridine A (1) and B (2). The pepteridines are produced in the pathogenic phenotypic variant and represent the first reported metabolites to be synthesized by a hybrid NRPS-pteridine pathway. These studies expand our view of the combinatorial biosynthetic potential available in bacteria.
Highlights
Nonribosomal peptides are a structurally and functionally privileged class of natural products constructed from a highly diverse pool of potential proteinogenic and nonproteinogenic amino acid building blocks (Walsh et al, 2013; Walsh, 2016)
Analogous carrier protein priming mechanisms have been proposed for branched chain fatty acid substrate utilization in the formation of N-acylamides and the pristinamycin IIa streptogramin antibiotic (Craig and Brady, 2011; Brachmann et al, 2012); and for a glycolicacyl-nonribosomal peptide synthetase (NRPS) extender unit in formation of the naphthyridinomycin antitumor antibiotic (Peng et al, 2012). In contrast to these pathways, we propose that the resulting loaded acyl-carrier protein in pepteridine biosynthesis would be unusually condensed with a free tetrahydropterin substrate by the atypical NRPS C domain to install the cisamide acyl-linkage
Bacterial natural products represent a rich source of lead structures in small molecule drug discovery efforts and serve as excellent molecular probes in biology
Summary
Nonribosomal peptides are a structurally and functionally privileged class of natural products constructed from a highly diverse pool of potential proteinogenic and nonproteinogenic amino acid building blocks (Walsh et al, 2013; Walsh, 2016). The core catalytic domains of a minimal nonribosomal peptide synthetase (NRPS) extender module include condensation (C), adenylation (A), and peptidyl-carrier protein (PCP, a.k.a., thiolation, T) domains. The NRPS first selects its cognate amino acid from the available substrate pool, a step controlled by the selectivity of the A domain, activates it as an aminoacyl adenylate, and subsequently loads it onto the PCP domain. The C domain typically catalyzes the formation of a trans-amide bond establishing individual peptide backbone linkages through nucleophilic attack of the free amino group present on a downstream aminoacyl-PCP on the upstream peptidyl-PCP. NRPSs can engage in ‘hybrid’ pathways to dramatically expand their biocatalytic capabilities
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